Abstract. LEXEO Therapeutics, LLC, is an early stage biotechnology company focused on using gene therapy technologies to protect vulnerable organs from oxidant stress. LEX01, the 1st LEXEO product, is a 1st in class, next generation gene therapy treatment for alpha 1-antitrypsin (AAT) deficiency, an autosomal recessive hereditary disorder associated with low serum levels of AAT. AAT (SERPINA1), a 52 kDa serine antiprotease produced by the liver, protects the lung from proteolytic enzymes released by inflammatory cells. In AAT deficiency, the lung is vulnerable to destruction by proteolytic enzymes released by inflammatory cells that degrade the lung, resulting in emphysema. AAT deficiency is currently treated with weekly infusions of AAT purified from pooled human plasma. Gene therapy for AAT deficiency holds the promise that a single administration will generate sufficient amounts of AAT to protect the lung on a persistent basis, obviating weekly AAT protein therapy. First generation gene therapy strategies are under development to treat AAT deficiency using adenoassociated virus (AAV) gene transfer vectors to deliver the human AAT coding sequence in vivo by various routes. The Achilles heel of these strategies is the sensitivity of the AAT molecule to oxidants. The active site of human AAT includes Met358, with a 2 contribution by Met351. When oxidized (e.g., by cigarette smoke, inflammatory cell products, ambient pollution), AAT is rendered ineffective and cannot inhibit neutrophil elastase, its primary target. LEX01, designed as an AAV vector coding for an elastase-inhibiting, oxidation-resistant human AAT, solves this problem with substitution at Met358 and/or Met351 with Val or Leu to render the AAT molecule oxidation resistant, yet maintaining its function as an anti-elastase. This is a Fast Track application with the goal of being clinical trial-ready within 3 yr. Successful completion of these aims will make LEXEO attractive for biotech, pharma and/or venture investment. Phase I, Aim 1. Using the normal M1(A213) AAT coding sequence as a base, assess combinations of Met, Leu and Val at positions 351 and 358 to determine the optimal neutrophil elastase inhibiting, oxidation resistant form of AAT to use in LEX01. Phase I, Aim 2. Determine that intrapleural administration of LEX01 to experimental animals results in persistent, high levels of oxidation-resistant human AAT in serum and lung epithelial lining fluid. Phase I, Aim 3. Have a pre- IND meeting with the FDA regarding the LEX01 development plan. Phase II, Aim 1. Produce and validate GMP clinical grade LEX01 and demonstrate in experimental animals it is safe to use in a human trial. Phase II, Aim 2. Develop and validate the assays to be used in the clinical study. Submit to the FDA an IND permitting initiation of a phase I clinical study of LEX01 for AAT deficiency.